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miRNAs are small non-coding RNA molecules which serve as promising biomarkers due to their important roles in the development and progression of various cancer types. The detection of miRNAs is of vital importance to the early-stage diagnostics and prognostics of multiple diseases. However, traditional detection strategies have faced some challenges owing to the intrinsic characteristics of miRNAs including small size, short sequence length, low concentration level and high sequence homology in complex real samples. To overcome these challenges, we proposed a MXene-enhanced plasmonic biosensor for real-time and label-free detection of miRNA. By utilizing MXene nanomaterial which possesses unique characteristics including large surface area and strong carrier confinement abilities, we tuned the absorption of our plasmonic sensing substrate to reach a “zero-reflection” state and induced an extremely sharp phase change at the resonance angle. Combined with the sensing mechanism based on phase-induced lateral displacement measurement, this MXene-enhanced plasmonic biosensor can achieve a much superior sensing performance compared to traditional SPR devices. Based on this biosensing scheme, the ultrasensitive detection of target miRNA with a detection limit down to 10 fM has been successfully demonstrated. More importantly, single-base mismatched miRNA can be easily distinguished from the target miRNA according to the sensing signal. Furthermore, our plasmonic biosensor is capable of detecting miRNA in complex media such as 100 % human serum samples without compromising the detection sensitivity. This MXene-enhanced plasmonic sensing scheme has the ability of detecting miRNAs with extremely low concentration levels in complex surrounding media without the need of introducing extra labels or amplification tags, which holds great potential in various biological applications and clinical diagnostics.